Antiproliferative activity and caspase enhancement properties of Annona muricata leaves extract against colorectal cancer cells

Latar belakang: Prevalensi kanker kolorektal meningkat di Asia termasuk di Indonesia. Annona muricata secara tradisional digunakan untuk menjaga kesehatan dan akhir-akhir ini digunakan untuk pasien kanker. Tujuan penelitian ini untuk mengetahui efek antiproliferatif ekstrak daun A. muricata dan kemampuannya meningkatkan aktivitas caspase in vitro dan ex vivo. Metode: Tiga puluh pasien kanker kolorektal secara acak dikelompokkan untuk menerima 300 mg ekstrak daun A. muricata (n=15) atau plasebo (n=15), setiap hari selama 8 minggu. Setelah itu serum pasien kanker kolorektal dari kedua kelompok digunakan untuk penelitian ex vivo untuk mengetahui efek ekstrak terhadap aktivitas caspase-8 dan caspase-9 cell line kanker kolorektal. Efek antiproliferatif ekstrak air daun A. muricata dan fraksinya dievaluasi terhadap cell line kanker kolorektal (DLD-1 dan COLO 205) dengan alat tes mikrokultur berdasarkan pengurangan metabolisme 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), dibandingkan dengan 5-fluorourasil dengan dosis 62,5- 2.000 μg/mL atau dengan plasebo. Data diuji dengan uji U Mann-Whitney. Batas kemaknaan ditetapkan pada nilai p≤0,05. Hasil: Fraksi larut etanol dari ekstrak air daun A. muricata menunjukkan efek sitotoksik pada sel kanker kolorektal DLD- 1 dan COLO 205, dengan konsentrasi hambatan lebih rendah dibanding plasebo dan 5-fluorourasil, masing-masing 20,59 μg/mL dan 654,9 μg/mL. Serum pasien yang minum ekstrak, bermakna dalam menstimulasi aktivitas caspase 9 (p=0,001) pada DLD-1 cell line kanker kolorektal, namun tidak bermakna dalam menstimulasi aktivitas caspase 8 (p=0,372). Kesimpulan: Hasil penelitian menunjukkan potensi sitotoksisitas ekstrak daun A. muricata dalam penelitian in vitro dan ex vivo kanker kolorektal. Background: The prevalence of colorectal cancer is rising in Asia including Indonesia. Annona muricata tea leaves, that is traditionally used for maintaining health, and lately being used by cancer patients. The objectives of this study is to investigate its effects in human colorectal cancer cell in vitro and ex vivo. Methods: Thirty patients with colorectal cancer (CRC) were enrolled in a randomized double-blind placebo-controlled trial. They were equally divided into two groups: those treated with 300 mg A. muricata leaf extract and placebo daily for 8 weeks. Serum from supplemented CRC patients of both groups was compared for caspase 9 and caspase 8 enhancement activity. Antiproliferative effect of water extract of A. muricata leaves and its fractions were evaluated against colorectal cancer cell line (DLD-1 and COLO 205) compared with 5-fluorouracil and placebo, the dose range was 62.5- 2,000 μg/mL. Method used was 3-(4,5-dimethylthiazol- 2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Data were analyzed by Mann-Whitney U test. The p value was set at 0.05. Results: Ethanol-soluble fraction of A. muricata leaves extract water extract (ESFAM) leaves extract had cytotoxicity effects on DLD-1 as well as COLO 205 cell line, as shown by the lower IC50 compared to 5-fluorouracil and placebo, 20.59 μg/mL and 654.9μg/mL, respectively. Serum of subjects supplemented with extract significantly induced caspase 9 (p=0.001) activity of DLD-1 colorectal cancer cell line, but not for caspase 8 activity (p=0.372). Conclusion: The studyʹs results suggest the cytotoxicity potential of A. muricata leaves extract in in vitro and ex vivo studies.